Soluble receptors, such as soluble TNFR (Enbrel®), have proven great value for therapeutical applications. Soluble receptors may also be used for diagnostic purposes, as well, e.g., for the screening of ligand-expression on diseased tissues, such as tumor-tissues in cancer patients.
The CD94/NKG2 family of receptors is composed of heterodimeric receptor members with activating or inhibitory potential. These receptors are expressed predominantly on NK cells and a subset of CD8+T cells, and they have been shown to play an important role in regulating responses against infected and tumorigenic cells. The main ligand for the CD94/NKG2 receptor is HLA-E. U.S. Pat. No. 6,262,244 to Houchins et al. described the human NKG2A sequence (SEQ ID NO:1), and Chang et al. (Eur J Immunol. 1995;25:2433-7) reported the human CD94 sequence (SEQ ID NO:2).
Soluble versions of CD94/NKG2 receptors are of interest not only as research tools but also as therapeutic agents. Both therapeutical and diagnostic applications, however, require stable soluble receptors that can be produced efficiently in a suitable biosystem. For many heterodimeric receptors such as, e.g., CD94/NKG2, this has, so far, proven difficult. Potential reasons are homo-dimerization of the single subunits, and that the more complex structure of heterodimeric receptors makes it more difficult to design soluble versions that are sufficiently stable.
Certain soluble CD94/NKG2A constructs, based on the expression of tagged soluble portions of the CD94 and NKG2A proteins, have been proposed in the literature (Brooks et al., J Immunol 1999:162:305-13; Ding et al Scand. J. Immunol. 1999;49:459-465; and Kaiser et al. Journal of Immunology, 2005, 174: 2878-2884). Soluble versions of other multimeric receptors, some of which fused to immunoglobulin Fc portions, have been described in, e.g., WO9937772, WO200208272, and WO20023237, relating to multimeric IL-18 receptor molecules; Wu et al. (Protein Sci. 1999;8:482-9), describing soluble forms of the IL-2 receptor; WO200222153, describing a soluble IL-20 receptor; WO200212345, relating to soluble ZCY-TOR 11 cytokine receptors, WO2002101006, relating to heteromultimeric proteins such as T-cell receptors, WO9533059, describing a heterodimeric receptor of gp130 and oncostatin M receptor beta chain, and U.S. Pat. No. 6,238,890, describing soluble forms of various glycoproteins. Soluble versions of heterodimeric T-cell receptors or subunits thereof have also been described in, e.g., Clements et al., Acta Crystallogr D Biol Crystallogr. 2002;58:21314; Laugel et al., J. Biol. Chem. 2005;280:1882-1892; Kim et al., J Mol Biol. 2000;302:899-916.
Further, U.S. Patent Publication Nos. 20030195338 and 20040072256 describe various types of Fc-linked receptor sequence proteins, U.S. Pat. No. 6,018,026 relates to dimerized fusion proteins; U.S. Pat. No. 6,833,441 relates to methods for generating chimeric heteromultimers; WO92/06204 and WO03012069 relate to, e.g., techniques for producing libraries of heterodimeric receptors; U.S. Patent Publication No. 20040138417 describes heteromultimer adhesins. Also, principles to promote heteromultimer formation are described in U.S. Patent Publication No. 20030078385, and reviewed in Marvin and Zhu, Acta Pharmacol Sin 2005;26(6):649-658 (see also Kontermann, Acta Pharmacol. Sin. 2005;26:1-9.
However, there remains a need for soluble versions of heterodimeric receptors such as CD94/NKG2 for diagnostic or therapeutic applications, and efficient methods of producing stable soluble heterodimeric receptors. The present invention addresses these and other needs in the art.